The fluoroquinolones represent a major class of antibacterials with great therapeutic potential. Over the years, several structure-activity and side-effect relationships have been developed, covering thousands of analogues, in an effort to improve overall antimicrobial efficacy while reducing undesirable side-effects. In this review, the various structural features of the quinolones which govern antibacterial efficacy and influence the side-effect profile are delineated and summarized at the molecular level. Those features which most remarkably enhance antimicrobial effectiveness are: a halogen (F or Cl) at the 8-position which improves oral absorption and activity against anaerobes; an alkylated pyrrolidine or piperazine at C7 which increases serum half-life and potency vs Gram-positive bacteria; and a cyclopropyl group at N1 and an amino substituent at C5, both of which improve overall potency. Some side-effects of the quinolones are class effects, and cannot be modulated by molecular variation. These include gastrointestinal irritation and arthropathy. Several other potential side-effects are directly influenced by structural modification. For example, CNS effects and drug interactions with theophylline and NSAIDs are strongly influenced by the C7 substituent with simple pyrrolidines and piperazines the worst actors. Increasing steric bulk through alkylation ameliorates these effects. Phototoxicity is determined by the nature of the 8-position substituent with halogen causing the greatest photo reaction while hydrogen and methoxy show little light induced toxicity. Genetic toxicity is controlled in additive fashion by the choice of groups at the 1, 7 and 8 positions. From the analysis, those groups which mutually improve efficacy while reducing side-effects are identified. In addition, preclinical models for determining potential side-effects are discussed.

Strategies for the treatment of human immunodeficiency virus-type 1 (HIV-1) infection must contend with the obstacle of drug resistance. HIV-1 nucleocapsid protein zinc fingers are prime antiviral targets because they are mutationally intolerant and are required both for acute infection and virion assembly. Nontoxic disulfide-substituted benzamides were identified that attack the zinc fingers, inactivate cell-free virions, inhibit acute and chronic infections, and exhibit broad antiretroviral activity. The compounds were highly synergistic with other antiviral agents, and resistant mutants have not been detected. Zinc finger-reactive compounds may offer an anti-HIV strategy that restricts drug-resistance development.

Fluoroquinolones are antibacterial agents that attack DNA gyrase and topoisomerase IV on chromosomal DNA. The existence of two f luoroquinolone targets and stepwise accumulation of resistance suggested that new quinolones could be found that would require cells to obtain two topoisomerase mutations to display resistance. For wild-type cells to become resistant, the two mutations must be acquired concomitantly. That is expected to occur infrequently. To identify such compounds, f luoroquinolones were tested for the ability to kill a moderately resistant gyrase mutant. Compounds containing a C8-methoxyl group were particularly lethal, and incubation of wild-type cultures on agar containing C8-methoxyl f luoroquinolones produced no resistant mutant, whereas thousands arose during comparable treatment with control compounds lacking the C8 substituent. When the test strain contained a preexisting topoisomerase IV mutation, which by itself conferred no resistance, equally high numbers of resistant mutants were obtained for C8-methoxyl and control compounds. Thus C8-methoxyl f luoroquinolones required two mutations for expression of resistance. Although highly lethal, C8-methoxyl f luoroquinolones were not more effective than C8-H controls at blocking bacterial growth. Consequently, quinolone action involves two events, which we envision as formation of drug-enzyme-DNA complexes followed by release of lethal double-strand DNA breaks. Release of DNA breaks, which must occur less frequently than complex formation, is probably the process stimulated by the C8-methoxyl group. Understanding this stimulation should provide insight into intracellular quinolone action and contribute to development of f luoroquinolones that prevent selection of resistant bacteria.

When Mycobacterium bovis BCG and Staphylococcus aureus were plated on agar containing increasing concentrations of fluoroquinolone, colony numbers exhibited a sharp drop, followed by a plateau and a second sharp drop. The plateau region correlated with the presence of first-step resistant mutants. Mutants were not recovered at concentrations above those required for the second sharp drop, thereby defining a mutant prevention concentration (MPC). A C-8-methoxy group lowered the MPC for anN-1-cyclopropyl fluoroquinolone.

A series of 60 newly synthesized and known quinolone antibacterials, including quinoline- and 1,8-naphthyridine-3-carboxylic acids, pyrido[2,3-d]pyrimidine-6-carboxylic acids, and some monocyclic 4-pyridone-3-carboxylic acids, were tested and compared in a newly established, easy to perform, DNA gyrase assay. The results were correlated with minimum inhibitory concentrations (MICs) against a variety of organisms. Among the known quinolones were 14 clinically significant drugs (oxolinic acid, norfloxacin, ciprofloxacin, enoxacin, etc.) which were used as standards and compared side-by-side. The study focused on the changes in DNA gyrase inhibition brought about by certain features of the molecules, namely, the C6-fluorine or the nature of the C7 substituent. The intrinsic gyrase inhibition of the fused parent rings, quinoline vs. naphthyridine vs. pyrido[2,3-d]pyrimidine, was also explored. In all cases, loss of enzyme inhibition produced poor MICs, but some compounds with good DNA gyrase inhibition did not correspondingly inhibit bacterial growth. Possible explanations for this phenomena and the benefits of a DNA gyrase-MIC strategy for developing future structure-activity relationships are discussed.

To obtain a general framework for understanding selection of antibiotic-resistant mutants, allelic diversity was examined with about 600 fluoroquinolone-resistant mutants of mycobacteria. Selection at low fluoroquinolone concentration produced many low-level resistance mutants. Some of these contained mutations that conferred unselected antibiotic resistance; none contained alterations in the quinolone-resistance-determining region of the GyrA protein, the principal drug target. As selection pressure increased, a variety of GyrA variants became prevalent. High concentrations of antibiotic reduced the variety to a few types, and eventually a concentration was reached at which no mutant was recovered. That concentration defined a threshold for preventing the selection of resistance. The pattern of variants selected, which was also strongly influenced by antibiotic structure, readily explained the variants present in clinical isolates. Thus, resistance arises from selection of mutants whose identity depends on drug concentration and structure, both of which can be manipulated to restrict selection.

Several disulfide benzamides have been shown to possess wide-spectrum antiretroviral activity in cell culture at low micromolar to submicromolar concentrations, inhibiting human immunodeficiency virus (HIV) type 1 (HIV-1) clinical and drug-resistant strains along with HIV-2 and simian immunodeficiency virus [Rice, W. G., Supko, J. G., Malspeis, L., Buckheit, R. W., Jr., Clanton, D., Bu, M., Graham, L., Schaeffer, C. A., Turpin, J. A., Domagala, J., Gogliotti, R., Bader, J. P., Halliday, S. M., Coren, L., Sowder, R. C., II, Arthur, L. O. & Henderson, L. E. (1995) Science 270, 1194-1197]. Rice and coworkers have proposed that the compounds act by "attacking" the two zinc fingers of HIV nucleocapsid protein. Shown here is evidence that low micromolar concentrations of the anti-HIV disulfide benzamides eject zinc from HIV nucleocapsid protein (NCp7) in vitro, as monitored by the zinc-specific fluorescent probe N-(6-methoxy-8-quinoyl)-p-toluenesulfonamide (TSQ). Structurally similar disulfide benzamides that do not inhibit HIV-1 in culture do not eject zinc, nor do analogs of the antiviral compounds with the disulfide replaced with a methylene sulfide. The kinetics of NCp7 zinc ejection by disulfide benzamides were found to be nonsaturable and biexponential, with the rate of ejection from the C-terminal zinc finger 7-fold faster than that from the N-terminal. The antiviral compounds were found to inhibit the zinc-dependent binding of NCp7 to HIV psi RNA, as studied by gel-shift assays, and the data correlated well with the zinc ejection data. Anti-HIV disulfide benzamides specifically eject NCp7 zinc and abolish the protein's ability to bind psi RNA in vitro, providing evidence for a possible antiretroviral mechanism of action of these compounds. Congeners of this class are under advanced preclinical evaluation as a potential chemotherapy for acquired immunodeficiency syndrome.

A series of 7,8-disubstituted 1-cyclopropyl-6-fluoroquinoline-3-carboxylic acids, 7-substituted 1-cyclopropyl-6-fluoro-1,8-naphthyridine-3-carboxylic acids, and 10-substituted 9-fluoropyridobenzoxazine-6-carboxylic acids has been prepared and evaluated for antibacterial activity. The side chains examined at the 7-position (benzoxazine 10-position) included piperazinyl (g), 3-aminopyrrolidinyl (a), 3-(aminomethyl)pyrrolidinyl (b), and alkylated 3-(aminomethyl)pyrrolidinyl (c-f). Variations at C-8 of the quinolone ring system included hydrogen, nitro, amino, fluorine, and chlorine. The relative enhancement of in vitro activities by the side chains on the 8-hydrogen quinolone and 1,8-naphthyridine against Gram-negative organisms was a greater than b greater than g greater than c-f. The activity imparted to the substituted quinolone nucleus by the 8-substituent was in the order F greater than Cl greater than naphthyridine greater than H greater than benzoxazine greater than NH2 greater than NO2. These trends were retained in vivo.